anatomy & connection types
Frontiers | Corticofugal GABAergic projection neurons in the mouse frontal cortex (frontiersin.org)
how to check projections of a inhibitory neuron from ofc to region-x
take gad67-gfp knock-in mouse(instead of gad67 which makes gaba, there is gfp). inject fast blue in region-x. inhibitory neurons that project from ofc to x, will be gfp and fb - double labelled(see page). later do ihc by using antibodies like anti-pv, anti-som to check for specific inhibitory neurons.
- https://elifesciences.org/articles/71103 - L2/3 connection types using 2p opto stimulation
How to check for both gfp and fb labelled neurons
- To check for neurons that are both Fast Blue (FB) and green fluorescent protein (GFP) labeled, you will need to use fluorescence microscopy to visualize the fluorescence of both tracers in the same tissue sample. Here is a general overview of the steps involved:
- Prepare the tissue sample: The tissue sample should be prepared using standard techniques for fluorescence microscopy. This usually involves fixing the tissue and sectioning it into thin slices. The slices can then be mounted on microscope slides and coverslipped.
- Visualize the FB signal: Use a fluorescence microscope to visualize the FB signal in the tissue. FB emits blue fluorescence when exposed to blue light, so you will need to use a filter that allows blue light to pass through while blocking other wavelengths of light. This will allow you to see the blue fluorescence of FB in the tissue.
- Visualize the GFP signal: Switch to a filter that allows green light to pass through while blocking other wavelengths of light. This will allow you to visualize the green fluorescence of GFP in the tissue.
- Overlay the images: To check for neurons that are both FB and GFP labeled, you will need to overlay the two images. You can do this using software that is designed for image processing and analysis. In the overlaid image, neurons that appear as both blue and green are likely to be double-labeled with FB and GFP.
- Confirm double labeling: Finally, you should confirm that the double-labeled neurons are in fact neurons and not other types of cells, such as glial cells. This can be done by examining the morphology of the labeled cells using other staining techniques, such as Nissl staining or immunohistochemistry. Additionally, it may be helpful to examine the distribution of the double-labeled neurons and compare it to the known distribution of the neurons labeled with each tracer separately.
chatgpt response if red + green → yellow neurons
if mcherry is injected in GFP expressing neuron, will it show yellow color under microscope
No, if mCherry (a red fluorescent protein) is injected into a green fluorescent protein (GFP) expressing neuron, it will not show yellow color under a microscope.
This is because mCherry and GFP fluoresce at different wavelengths. GFP fluoresces in the green spectrum, while mCherry fluoresces in the red spectrum. Therefore, when both proteins are present in the same neuron, they will emit green and red fluorescence, respectively, and will not mix to produce yellow color.
However, it is possible to use imaging techniques such as spectral unmixing to separate the green and red signals and analyze them separately. This technique can help distinguish different subcellular structures or processes that are labeled with GFP and mCherry in the same neuron.
Regenerate response